In this proposal we intend to examine several mutant strains of Escherichia coli which are hypersensitive to the ribosomal antibiotics streptomycin, spectinomycin or erythromycin. These mutants are also temperature-sensitive for some aspect of protein synthesis. Our initial studies will involve using genetic methods to examine the relationship between the antibiotic hypersensitive and temperature-sensitive phenotypes of these mutants. We will examine the translational activity of the ribosomes from these cells to see if the hypersensitivity is an intrinsic characteristic of these particles. The fidelity of messenger RNA translation by these ribosomes will also be studied. We also intend to look at antibiotic binding to the subunits of the ribosomes. Finally, we propose to look for ribosomal protein alterations in these mutant strains, to help localize the site of the altered antibiotic sensitivity. The results from these investigations should give some new insights into the mechanism of antibiotic inhibition of protein synthesis. We hope to give a better definition of the specific components of the translational machinery which are the targets for inhibition. An increased understanding of the inhibitory action of ribosomal antibiotics is important for the effective use of these compounds in controlling infection.